Summary- Electrical excitability and Ca2+ influx are key features of neurons, endocrine cells, and cardiovascular muscle. However, excessive Ca2+ influx can sensitize these cells to cytotoxic agents, and activate pathological cellular events. Ca influx via Cav1.3 L-type Ca channels is hypothesized to play a key 2+ 2+ role in the death of dopaminergic neurons in Parkinson's Disease, and in cytokine-mediated cell death in pancreatic beta cells during the onset of Type 1 diabetes. Therefore, the development of Cav1.3 inhibitors as potential treatments for Parkinson's disease and diabetes is being pursued. Current drugs that inhibit Cav1.3 also block the closely related channel Cav1.2. Selective Cav1.3 inhibitors would be desirable since they could potentially protect neurons and endocrine cells without inducing hypotension. We've found that the 60 amino acid extracellular IIIS5-3P loops of these channels confer distinct pharmacological properties, and shown that the isolated Cav1.2 IIIS5-3P loop can bind the toxin calcicludine using biolayer interferometry (BLI). In Aim 1, we'll screen a 2.5 x 10 membered, DNA-coded chemical library for binding to the IIIS5-3P loop of Cav1.2 or 8 Cav1.3., confirm binding using BLI, and assess the hits for the ability to selectively inhibit Cav1.2 or Cav1.3 activity using whole-cell electrophysiology. We've also found that the intracellular II-III loop of Cav1.3 selectively enhances inactivation of Cav1.3 but not that of Cav1.2 or a C-terminal truncated splice variant of Cav1.3. In Aim 2, we'll define the minimal peptide sequence within the Cav1.3 II-III loop that confers this activity, and define the role of auxiliary beta subunits and the C-terminal tail of Cav1.3 in the mechanism of action. Completion of these aims will provide structural templates for the development of Cav1.2-selective inhibitors, and two mechanistically distinct classes of Cav1.3-selective inhibitors.